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Thyroid dysfunction has been implicated as a potential pathophysiological factor in glucose homeostasis and insulin resistance (IR). This study aimed to identify the correlation between thyroid dysfunction and IR. We used data from the sixth Korean National Health and Nutrition Examination Survey to evaluate a total of 5727 participants. The triglyceride glucose (TyG) index and homeostasis model assessment of insulin resistance (HOMA-IR) were calculated to represent IR. Correlation analysis was performed between thyroid dysfunction and IR. The log-transformed TSH (LnTSH) and free T4 were significantly correlated with the TyG index (TSH, beta coefficient 0.025, 95% confidence interval [CI] 0.014–0.036, p p p p p

An analytical method was validated and developed to determine the levels of volatile organic compounds (VOCs) including 1,3-butadiene, isoprene, acrylonitrile, benzene, toluene, and styrene from smoking cessation aids using by gas chromatography-mass spectrometry (GC-MS). Seventeen non-ignitable cigarette type of smoking cessation aids, liquid-phase of smoking cessation aids, gas-phase of smoking cessation aids, and ignitable cigarette type of smoking cessation aids were analyzed for levels of six VOCs. The proposed method for six VOCs was validated in satisfactory linearity (R

This study analyzed four polycyclic aromatic hydrocarbons (PAHs) from frequently consumed home meal replacement (HMR) products in Korea and evaluated their chemical analysis methods. The PAHs investigated were benzo[a]anthracene, chrysene, benzo[b]fluoranthene and benzo[a]pyrene. Chrysene-d12 and benzo[a]pyrene-d12 served as internal standards. The sample was dissolved in dichloromethane before extraction. Liquid–liquid extraction, microwave extraction, alkali digestion and GC–MS were used for analysis. Method validation was conducted on four matrices: fatty solid, fatty liquid, non-fatty liquid, non-fatty solid. Linear correlation coefficients (R(2)) were all above 0.99 and accuracy ranged from 80.03 to 119.65%. The LOD and LOQ were in the range of 0.03–0.15 and 0.09–0.44 μg/kg, respectively. The recoveries varied from 81.09 to 116.42% and precision ranged from 0.07 to 10.73% in both intraday and interday analysis. All concentrations of total PAHs from HMR products were detected at relatively low concentration. This study could provide the PAHs content from HMR products in Korea.

Molten oxide electrolysis can be used to reduce the carbon dioxide emissions from ironmaking. In this paper, we electrochemically decompose iron oxide (Fe2O3) into iron (Fe) and oxygen (O2) using b...

Type 2 diabetes mellitus (T2DM) is the leading cause of chronic kidney disease (CKD) and almost 40% of T2DM patients develop diabetic kidney disease (DKD), characterized by reduction in estimated glomerular filtration rate (eGFR) and/or increased albuminuria. The patients with DKD are at increased risk of cardiovascular disease (CVD) and cardiovascular (CV) death, therefore renal protection strategies are very important in T2DM patients. Renin-angiotensin-aldosterone system (RAAS) inhibitors are well known, classic drugs for DKD therapy. Recently, several clinical trials have reported some newer glucose-lowering drugs, including sodium-glucose cotransporter 2 (SGLT2) inhibitors and glucagon-like peptide-1 receptor agonists (GLP-1RAs), have beneficial effect on kidney in T2DM patients. Currently, there are five GLP-1RAs (exenatide, lixisenatide, liraglutide, dulaglutide, and semaglutide) approved in Europe and USA. All GLP-1RAs have shown the CV benefits and safety. Most of these trials have the data of secondary exploratory renal endpoints, and the secondary renal outcomes from GLP-1RA CV outcome trials suggest a renal protective effect of GLP-1RA1. Based on the results of the previous trials, GLP-1RAs have been recommended in DKD patients if they are contraindicated or not tolerated to SGLT2 inhibitors.

Osteoporosis is a disease in which bone mineral density decreases due to abnormal activity of osteoclasts, and is commonly found in post-menopausal women who have decreased levels of female hormones. Sphingosylphosphorylcholine (SPC) is an important biological lipid that can be converted to sphingosine-1-phosphate (S1P) by autotaxin. S1P is known to be involved in osteoclast activation by stimulating osteoblasts, but bone regulation by SPC is not well understood. In this study, we found that SPC strongly inhibits RANKL-induced osteoclast differentiation. SPC-induced inhibitory effects on osteoclast differentiation were not affected by several antagonists of S1P receptors or pertussis toxin, suggesting cell surface receptor independency. However, SPC inhibited RANKL-induced calcineurin activation and subsequent NFATc1 activity, leading to decrease of the expression of Trap and Ctsk. Moreover, we found that bone loss in an experimental osteoporosis mouse model was recovered by SPC injection. SPC also blocked ovariectomy-induced body weight increase and Nfatc1 gene expression in mice. We also found that SPC inhibits RANKL-induced osteoclast differentiation in human macrophages. Since currently available treatments for osteoporosis, such as administration of female hormones or hormone receptor modulators, show serious side effects, SPC has potential as a new agent for osteoporosis treatment.

Codonopsis lanceolata has been widely used as an anti-inflammatory and anti-lipogenic agent in traditional medicine. Recently, C. lanceolata was reported to prevent hypertension by improving vascular function. This study evaluated the effects of C. lanceolata and its major component lancemaside A on cytosolic calcium concentration in vascular endothelial cells and vascular smooth muscle cells. Cytosolic calcium concentration was measured using fura-2 AM fluorescence. C. lanceolata or lancemaside A increased the cytosolic calcium concentration by releasing Ca2+ from the endoplasmic reticulum and sarcoplasmic reticulum and by Ca2+ entry into endothelial cells and vascular smooth muscle cells from extracellular sources. The C. lanceolata- and lancemaside A-induced cytosolic calcium concentration increases were significantly inhibited by lanthanum, an inhibitor of non-selective cation channels, in both endothelial cells and vascular smooth muscle cells. Moreover, C. lanceolata and lancemaside A significantly inhibited store-operated Ca2+ entry under pathological extracellular Ca2+ levels. In Ca2+-free extracellular fluid, increases in the cytosolic calcium concentration induced by C. lanceolata or lancemaside A were significantly inhibited by U73122, an inhibitor of phospholipase C, and 2-APB, an inositol 1,4,5-trisphosphate receptor antagonist. In addition, dantrolene treatment, which inhibits Ca2+ release through ryanodine receptor channels, also inhibited C. lanceolata- or lancemaside A-induced increases in the cytosolic calcium concentration through the phospholipase C/inositol 1,4,5-trisphosphate pathway. These results suggest that C. lanceolata and lancemaside A increase the cytosolic calcium concentration through the non-selective cation channels and phospholipase C/inositol 1,4,5-trisphosphate pathways under physiological conditions and inhibit store-operated Ca2+ entry under pathological conditions in endothelial cells and vascular smooth muscle cells. C. lanceolata or lancemaside A can protect endothelial cells and vascular smooth muscle cells by maintaining cytosolic calcium concentration homeostasis, suggesting possible applications for these materials in diets for preventing vascular damage.

Owing to the limitations of conventional cancer therapies, cancer immunotherapy has emerged for the prevention of cancer recurrence. To provoke adaptive immune responses that are antigen-specific, it is important to develop an efficient antigen delivery system that can enhance the activation and maturation of the dendritic cells (DCs) in the human body. In this study, we synthesize hollow mesoporous silica nanoparticles with extra-large mesopores (H-XL-MSNs) based on a single-step synthesis from core-shell mesoporous silica nanoparticles with a core composed of an assembly of iron oxide nanoparticles. The hollow void inside the mesoporous silica nanoparticles with large mesopores allows a high loading efficiency of various model proteins of different sizes. The H-XL-MSNs are coated with a poly(ethyleneimine) (PEI) solution to provide an immune adjuvant and change the surface charge of the particles for loading and slow release of a model antigen. An in vitro study using a cancer vaccine based on the PEI-coated H-XL-MSNs with the loading of the model antigen showed an enhanced activation of the DCs. An in vivo study demonstrated that the resulting cancer vaccine increased the antigen-specific cytotoxic T cells, enhanced the suppression of tumor growth, and improved the survival rate after challenging cancer to mice. These findings suggest that these hollow MSNs with extra-large pores can be used as excellent antigen carriers for immunotherapy.

The inherent conformational preferences of the neutral β-peptide foldamer series, Ac-(ACHC)n-NHBn, n = 2-4, are studied in the gas phase using conformation-specific IR-UV double resonance methods. The cyclically constrained chiral β-amino acid cis-2-aminocyclohexane carboxylic acid (ACHC) is designed to bring both right- and left-handed helices into close energetic proximity. Comparison of the infrared spectra in the NH stretch and amide I/II regions with the predictions of DFT calculations lead to the unambiguous assignment of four out of the six observed conformations of the molecules in this series, while corroborating computational and spectral evidence, affords tentative assignments of the remaining two conformers for which IR data were not recorded. The observed structures fall into one of two conformational families: a right-handed 12/10-mixed helix or its "cap-disrupted" left-handed helical analogue, which coexist with significant populations. Site-specific and stereospecific methylation on the cyclohexane backbone at the dipeptide (n = 2) level is also tested as a means to sterically lock in a predetermined cyclohexane chair conformation. These substitutions are proven to be a means of selectively driving formation of one helical screw sense or the other. Calculated relative energies and free energies of all possible structures for the molecules provide strong supporting evidence that the rigid nature of the ACHC residue confers unusual stability to the 12/10-mixed helix conformation, regardless of local environment, temperature, or C-terminal capping unit. The simultaneous presence of both handed helices offers unique opportunities for future studies of their interconversion.

The ClpP serine peptidase is a tetradecameric degradation machine involved in many physiological processes. It becomes a competent ATP-dependent protease with Clp-ATPases. Small chemical compounds, acyldepsipeptides (ADEPs), are known to cause dysregulation and activation of ClpP without ATPases, and have potential as novel antibiotics. Previously, structural studies of ClpP from various species revealed the structural details, conformational changes, and activation mechanism. Although product release by the side exit pores has been proposed, the detailed driving force for product release remains elusive. Here, we report crystal structures of ClpP from Bacillus subtilis (BsClpP) in unforeseen ADEP-bound states. Cryo-electron microscopy structures revealed various conformational states at different pH conditions. To understand the conformational change for product release, we investigated the relationship between substrate hydrolysis and the pH lowering process. Our data, together with previous findings, provide insight into the molecular mechanism of product release by ClpP self-compartmentalizing protease.

One of the well-known causes of hearing loss is noise. Approximately 31.1% of Americans between the ages of 20 and 69 years (61.1 million people) have high-frequency hearing loss associated with noise exposure. In addition, recurrent noise exposure can accelerate age-related hearing loss. Phlorofucofuroeckol A (PFF-A) and dieckol, polyphenols extracted from the brown alga Ecklonia cava, are potent antioxidant agents. In this study, we investigated the effect of PFF-A and dieckol on the consequences of noise exposure in mice. In 1,1-diphenyl-2-picrylhydrazyl assay, dieckol and PFF-A both showed significant radical-scavenging activity. The mice were exposed to 115 dB SPL of noise one single time for 2 h. Auditory brainstem response(ABR) threshold shifts 4 h after 4 kHz noise exposure in mice that received dieckol were significantly lower than those in the saline with noise group. The high-PFF-A group showed a lower threshold shift at click and 16 kHz 1 day after noise exposure than the control group. The high-PFF-A group also showed higher hair cell survival than in the control at 3 days after exposure in the apical turn. These results suggest that noise-induced hair cell damage in cochlear and the ABR threshold shift can be alleviated by dieckol and PFF-A in the mouse. Derivatives of these compounds may be applied to individuals who are inevitably exposed to noise, contributing to the prevention of noise-induced hearing loss with a low probability of adverse effects.

New bone anabolic agents for the effective treatment of bone metabolic diseases like osteoporosis are of high clinical demand. In the present study, we reveal the function of salt-inducible kinase 1 (SIK1) in regulating osteoblast differentiation. Gene knockdown of SIK1 but not of SIK2 or SIK3 expression in primary preosteoblasts increased osteoblast differentiation and bone matrix mineralization. SIK1 also regulated the proliferation of osteoblastic precursor cells in osteogenesis. This negative control of osteoblasts required the catalytic activity of SIK1. SIK1 phosphorylated CREB regulated transcription coactivator 1 (CRTC1), preventing CRTC1 from enhancing CREB transcriptional activity for the expression of osteogenic genes like Id1. Furthermore, SIK1 knockout (KO) mice had higher bone mass, osteoblast number, and bone formation rate versus littermate wild-type (WT) mice. Preosteoblasts from SIK1 KO mice showed more osteoblastogenic potential than did WT cells, whereas osteoclast generation among KO and WT precursors was indifferent. In addition, bone morphogenic protein 2 (BMP2) suppressed both SIK1 expression as well as SIK1 activity by protein kinase A (PKA)–dependent mechanisms to stimulate osteogenesis. Taken together, our results indicate that SIK1 is a key negative regulator of preosteoblast proliferation and osteoblast differentiation and that the repression of SIK1 is crucial for BMP2 signaling for osteogenesis. Therefore, we propose SIK1 to be a useful therapeutic target for the development of bone anabolic strategies.

Mitofusin 2 (Mfn2) is a mitochondrial outer membrane protein that participates in tethering mitochondria to the ER. Mitochondria-ER tethering has been demonstrated to play important roles in many cellular activities by regulating homeostasis of metabolites and calcium. Intracellular calcium signaling is crucial for the differentiation of osteoclasts, the bone-resorbing cells. In this study, we investigated whether Mfn2 plays a role in osteoclastogenesis by receptor activator of nuclear factor kappa B (RANKL) in primary cells. We found that RANKL increased Mfn2 expression during osteoclast formation from mouse bone marrow-derived macrophages (BMMs). When Mfn2 expression was suppressed in BMMs by using a siRNA-mediated gene knock-down system, osteoclast differentiation and activity of mature osteoclasts were reduced. Mfn2 knock-down also decreased the RANKL-mediated induction of NFATc1, the key transcription factor for osteoclast gene expression, without affecting c-Fos level. This effect on NFATc1 was associated with decreased calcium oscillation and calcineurin activity in Mfn2-deficient osteoclasts. Taken together, our results indicate that Mfn2 positively contributes to RANKL-induced osteoclast differentiation by regulating the calcium-calcieurin-NFATc1 axis, raising the importance of a previously under-recognized role of mitochondria in osteoclastogenesis.

Rates of microbial drug resistance are increasing worldwide; therefore, antimicrobial peptides (AMPs) are considered promising alternative therapeutic agents to antibiotics. AMPs are essential components of the innate immune system and exhibit broad-spectrum antimicrobial activity. P5 is a Cecropin A-Magainin 2 hybrid analog peptide with antimicrobial activity against Gram-negative and Gram-positive bacteria. In the present study, truncated peptides were designed to reduction length, retainment their antimicrobial activity and low toxicity at high concentrations compared with that of the parent peptide P5. The truncated peptides P5-CT1 and P5-NT1 exhibited antibacterial activities against both Gram-negative and Gram-positive bacteria. In contrast, P5-CT2, P5-CT3, P5-NT2, and P5-NT3 showed higher antibacterial activities against gram-positive bacteria compared to Gram-negative bacteria at low concentration of peptides. The truncated peptides showed lower hemolytic activity and toxic effects against mammalian cells compared with those of the parent peptide P5. The levels of several truncated peptides were maintained in the presence of physiological concentrations of salts, indicating their high stability. The results of flow cytometry, propidium iodide uptake, n-phenyl-1-naphthylamine uptake, and 3,3′-dipropylthiadicarbocyanine iodide assays showed that these truncated peptides killed microbial cells by increasing membrane permeability, thereby causing membrane damage. The results suggested that truncated peptides of P5 have good potential for use as novel antimicrobial agents.

We have identified a mechanism to diminish the proliferative capacity of cells during cell expansion using human adiposederived stromal cells (hAD-SCs) as a model of replicative senescence. hAD-SCs of high-passage numbers exhibited a reduced proliferative capacity with accelerated cellular senescence. Levels of key bioactive sphingolipids were significantly increased in these senescent hAD-SCs. Notably, the transcription of sphingosine kinase 1 (SPHK1) was down-regulated in hAD-SCs at high-passage numbers. SPHK1 knockdown as well as inhibition of its enzymatic activity impeded the proliferation of hAD-SCs, with concomitant induction of cellular senescence and accumulation of sphingolipids, as seen in high-passage cells. SPHK1 knockdown-accelerated cellular senescence was attenuated by co-treatment with sphingosine-1-phosphate and an inhibitor of ceramide synthesis, fumonisin B1, but not by treatment with either one alone. Together, these results suggest that transcriptional down-regulation of SPHK1 is a critical inducer of altered sphingolipid profiles and enhances replicative senescence during multiple rounds of cell division. [BMB Reports 2019; 52(3): 220-225].

An amphipathic α-helical peptide, Hp1404, was isolated from the venomous gland of the scorpion Heterometrus petersii. Hp1404 exhibits antimicrobial activity against methicillin-resistant Staphylococcus aureus but is cytotoxic. In this study, we designed antimicrobial peptides by substituting amino acids at the 14 C-terminal residues of Hp1404 to reduce toxicity and improve antibacterial activity. The analog peptides, which had an amphipathic α-helical structure, were active against gram-positive and gram-negative bacteria, particularly multidrug-resistant Acinetobacter baumannii, and showed lower cytotoxicity than Hp1404. N-phenyl-1-naphthylamine uptake and DisC3-5 assays demonstrated that the peptides kill bacteria by effectively permeating the outer and cytoplasmic membranes. Additionally, the analog peptides inhibited biofilm formation largely than Hp1404 at low concentrations. These results suggest that the analog peptides of Hp1404 can be used as therapeutic agents against A. baumannii infection.

Background Exposure to ozone (O3) induces neutrophilic inflammation and goblet cell hyperplasia in humans and experimental animals. Because the solute carrier family 26-member 4 (Slc26a4; pendrin) gene induces mucin production and intraluminal acidification in the airways, it was hypothesized to be a key molecule in O3-induced airway injury. Thus, we evaluated the role of Slc26a4 and the protective effects of ammonium chloride (NH4Cl) in O3-induced airway injury in mice. Methods Six-week-old female BALB/c mice were exposed to filtered air or O3 for 21 days (2 ppm for 3 hr/day). NH4Cl (0, 0.1, 1, and 10 mM) was administered intratracheally into the airways. Airway resistance was measured using a flexiVent system, and bronchoalveolar lavage fluid (BALF) cells were differentially counted. Slc26a4 and Muc5ac proteins and mRNA were measured via western blotting, real-time polymerase chain reaction, and immunostaining. Tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-17, IL-1β, and caspase-1 were analyzed via western blotting. Results The levels Slc26a4 protein and mRNA significantly increased in lung tissues from Day 7 to Day 21 of O3 exposure, with concomitant increases in lung resistance, numbers of goblet cells in lung tissues, and inflammatory cells and thiocyanate (SCN−) levels in BALF in a time-dependent manner. Treatment with NH4Cl significantly reduced these changes to levels similar to those of sham-treated mice, with a concomitant reduction of Slc26a4 proteins in lung lysates and SCN− levels in BALF. Slc26a4 protein was co-expressed with muc5ac protein in the bronchial epithelium, as indicated by immunofluorescence staining. NH4Cl treatment also significantly attenuated the O3-induced increases in IFN-γ, TNF-α, IL-17, IL-1β, and p20-activated caspase-1. Conclusion Slc26a4 may be involved in O3-induced inflammatory and epithelial changes in the airways via activation of the inflammasome and the induction of IL-17 and IFN-γ. NH4Cl shows a potential as a therapeutic agent for controlling O3-induced airway inflammation and epithelial damage by modulating Slc26a4 expression., Graphical Abstract

Atopic dermatitis (AD) is an inflammatory skin disease characterized by intense pruritus and relapsable eczematous lesions. The hallmarks of AD are defects in the epidermal barrier and immunoglobulin E (IgE)-mediated sensitization to several environmental allergens, as well as an immune disorder mediated by an imbalance toward T-helper-2 response. Melittin, a major component of bee venom, has been studied in various inflammatory diseases. However, the beneficial effects of melittin on mouse with AD-like symptoms have not been explored. Therefore, we investigated the anti-allergic effects of melittin. AD was induced by ovalbumin (OVA) patch. After agent treatment, skin tissues and sera were extracted from the sacrificed mice were used to demonstrate the effects of melittin through various molecular biological methods. The results showed that OVA-induced skin thickening and inflammatory infiltration were decreased in the melittin-treated group. Melittin prevented OVA-induced filaggrin deficiency and imbalanced inflammatory mediators. Furthermore, melittin inhibited IL-4/IL-13-induced filaggrin downregulation through the blockade of STAT3 activation in human keratinocytes. In summary, this study has shown that melittin ameliorated OVA-induced AD-like symptoms from various perspectives. The findings of this study may be the first evidence of the anti-inflammatory effects of melittin on OVA-induced AD.

Mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4) is implicated in type 2 diabetes mellitus, insulin tolerance, inflammation, cancer, and atherosclerosis. We found that GNE 495 and PF 06260933 (both potent and selective MAP4K4 inhibitors) regulated human platelet activation. Immunoblotting revealed human platelets express MAP4K4, and that GNE 495 and PF 06260933 inhibited collagen-, ADP-, and thrombin-induced platelet aggregation and eventually suppressed granule release, TXA2 generation, integrin αIIbβ3 activation, and clot retraction. In addition, both inhibitors elevated intracellular levels of cAMP, and coincubation with GNE 495 and aspirin or dipyridamole (a phosphodiesterase inhibitor) synergistically inhibited collagen-induced platelet aggregation and TXA2 generation. Moreover, both inhibitors phosphorylated VASP (ser157), IP3 receptor, and PKA and attenuated MAPK and PI3K/Akt/GSK3β signaling pathways. This study is the first to demonstrate that MAP4K4 inhibitors reduce thrombus formation by inhibiting platelet activation. These findings also suggest MAP4K4 be considered an emerging target protein for the treatment of thrombosis.

Inverse opal hydrogels (IOHs) for cell culture were fabricated and optimized using calcium-crosslinked alginate microbeads as sacrificial template and gelatin as a matrix. In contrast to traditional three-dimensional (3D) scaffolds, the gelatin IOHs allowed the utilization of both the macropore surface and inner matrix for cell co-culture. In order to remove templates efficiently for the construction of 3D interconnected macropores and to maintain high cell viability during the template removal process using EDTA solution, various factors in fabrication, including alginate viscosity, alginate concentration, alginate microbeads size, crosslinking calcium concentration, and gelatin network density were investigated. Low viscosity alginate, lower crosslinking calcium ion concentration, and lower concentration of alginate and gelatin were found to obtain high viability of cells encapsulated in the gelatin matrix after removal of the alginate template by EDTA treatment by allowing rapid dissociation and diffusion of alginate polymers. Based on the optimized fabrication conditions, gelatin IOHs showed good potential as a cell co-culture system, applicable to tissue engineering and cancer research.

We have constructed and experimentally tested a comprehensive genome-scale model of photoautotrophic growth, denoted iSyp821, for the cyanobacterium Synechococcus sp. PCC 7002. iSyp821 incorporates a variable biomass objective function (vBOF), in which stoichiometries of the major biomass components vary according to light intensity. The vBOF was constrained to fit the measured cellular carbohydrate/protein content under different light intensities. iSyp821 provides rigorous agreement with experimentally measured cell growth rates and inorganic carbon uptake rates as a function of light intensity. iSyp821 predicts two observed metabolic transitions that occur as light intensity increases: 1) from PSI-cyclic to linear electron flow (greater redox energy), and 2) from carbon allocation as proteins (growth) to carbohydrates (energy storage) mode. iSyp821 predicts photoautotrophic carbon flux into 1) a hybrid gluconeogenesis-pentose phosphate (PP) pathway that produces glycogen by an alternative pathway than conventional gluconeogenesis, and 2) the photorespiration pathway to synthesize the essential amino acid, glycine. Quantitative fluxes through both pathways were verified experimentally by following the kinetics of formation of 13C metabolites from 13CO2 fixation. iSyp821 was modified to include changes in gene products (enzymes) from experimentally measured transcriptomic data and applied to estimate changes in concentrations of metabolites arising from nutrient stress. Using this strategy, we found that iSyp821 correctly predicts the observed redistribution pattern of carbon products under nitrogen depletion, including decreased rates of CO2 uptake, amino acid synthesis, and increased rates of glycogen and lipid synthesis. Refereed/Peer-reviewed

Patients with inflammatory bone disease or cancer exhibit an increased risk of fractures and delayed bone healing. The S100A4 protein is a member of the calcium-binding S100 protein family, which is abundantly expressed in inflammatory diseases and cancers. We investigated the effects of extracellular S100A4 on osteoblasts, which are cells responsible for bone formation. Treating primary calvarial osteoblasts with recombinant S100A4 resulted in matrix mineralization reductions. The expression of osteoblast marker genes including osteocalcin and osterix was also suppressed. Interestingly, S100A4 stimulated the nuclear factor-kappaB (NF-κB) signaling pathway in osteoblasts. More importantly, the ex vivo organ culture of mouse calvariae with recombinant S100A4 decreased the expression levels of osteocalcin, supporting the results of our in vitro experiments. This suggests that extracellular S100A4 is important for the regulation of bone formation by activating the NF-κB signaling pathway in osteoblasts. [BMB Reports 2017; 50(2): 97-102].

Osteoclasts (OCs), cells specialized for bone resorption, are generated from monocyte/macrophage precursors by a differentiation process governed by RANKL. Here, we show that DCTN1, a key component of the dynactin complex, plays important roles in OC differentiation. The expression of DCTN1 was upregulated by RANKL. The inhibition of DCTN1 expression by gene knockdown suppressed OC formation, bone resorption, and the induction of NFATc1 and c-Fos, critical transcription factors for osteoclastogenesis. More importantly, the activation of Cdc42 by RANKL was inhibited upon DCTN1 silencing. The forced expression of constitutively active Cdc42 restored the OC differentiation of precursors with DCTN1 deletion. In addition, PAK2 was found to be activated by RANKL and to function downstream of Cdc42. The DCTN1-Cdc42 axis also inhibited apoptosis and caspase-3 activation. Furthermore, the anti-osteoclastogenic effect of DCTN1 knockdown was verified in an animal model of bone erosion. Intriguingly, DCTN1 overexpression was also detrimental to OC differentiation, suggesting that DCTN1 should be regulated at the appropriate level for effective osteoclastogenesis. Collectively, our results reveal that DCTN1 participates in the activation of Cdc42/PAK2 signaling and the inhibition of apoptosis during osteoclastogenesis., Bone remodeling: Regulatory protein identified A critical mechanism for maintaining bone health uncovered by scientists in South Korea could provide insights into bone disease development. Bone remodeling is a lifetime process of bone generation that ensures bones remain healthy. Osteoclasts (OC), cells that break down bone, differentiate from white blood cell populations. Disruption to OC formation and function plays a critical role in bone diseases, yet the regulatory mechanisms in OC generation are unclear. Hong-Hee Kim at Seoul National University and co-workers investigated the role of a protein called DCTN1, which is involved in skeletal assembly processes. The team found that inhibiting DCTN1 suppressed the expression of key proteins needed for OC formation in cell cultures and mouse models. Overexpressing DCTN1 was equally damaging, suggesting the protein plays a key regulatory role.

MotivationDetermining intracellular metabolic flux through isotope labeling techniques such as 13C metabolic flux analysis (13C-MFA) incurs significant cost and effort. Previous studies have shown transcriptomic data coupled with constraint-based metabolic modeling can determine intracellular fluxes that correlate highly with 13C-MFA measured fluxes and can achieve higher accuracy than constraint-based metabolic modeling alone. These studies, however, used validation data limited to E. coli and S. cerevisiae grown on glucose, with significantly similar flux distribution for central metabolism. It is unclear whether those results apply to more diverse metabolisms, and therefore further, extensive validation is needed.ResultsIn this paper, we formed a dataset of transcriptomic data coupled with corresponding 13C-MFA flux data for 21 experimental conditions in different unicellular organisms grown on varying carbon substrates and conditions. Three computational flux-balance analysis (FBA) methods were comparatively assessed. The results show when uptake rates of carbon sources and key metabolites are known, transcriptomic data provides no significant advantage over constraint-based metabolic modeling (average correlation coefficients, transcriptomic E-Flux2 0.725 and SPOT 0.650 vs non-transcriptomic pFBA 0.768). When uptake rates are unknown, however, predictions obtained utilizing transcriptomic data are generally good and significantly better than those obtained using constraint-based metabolic modeling alone (E-Flux2 0.385 and SPOT 0.583 vs pFBA 0.237). Thus, transcriptomic data coupled with constraint-based metabolic modeling is a promising method to obtain intracellular flux estimates in microorganisms, particularly in cases where uptake rates of key metabolites cannot be easily determined, such as for growth in complex media or in vivo conditions.

A polyene compound NPP identified in Pseudonocardia autotrophica was shown to contain an aglycone identical to nystatin, but to harbor a unique disaccharide moiety that led to higher solubility and reduced hemolytic activity. Recently, it was revealed that the final step of NPP (nystatin-like polyene) biosynthesis is C10 regio-specific hydroxylation by the cytochrome P450 hydroxylase (CYP) NppL (Kim et al. [7]). Through mutation and cross-complementation, here we found that NppL preferred a polyene substrate containing a disaccharide moiety for C10 hydroxylation, while its orthologue NysL involved in nystatin biosynthesis showed no substrate preference toward mono- and disaccharide moieties, suggesting that two homologous polyene CYPs, NppL and NysL might possess a unique domain recognizing a sugar moiety. Two hybrid NppL constructs containing the C-terminal domain of NysL exhibited no substrate preference toward 10-deoxy NPP and 10-deoxy nystatin-like NysL, implying that the C-terminal domain plays a major role in differentiating the sugar moiety responsible for substrate specificity. Further C-terminal domain dissection of NppL revealed that the last fifty amino acids play a critical role in determining substrate specificity of polyene-specific hydroxylation, setting the stage for the biotechnological application of hydroxyl diversification for novel polyene biosynthesis in actinomycetes.

Amorphous biodegradable copolyesters have been synthesized using various alicyclic monomers, including 1,4-cyclohexanedimethanol (CHDM), 1,4-cyclohexanediol (CHDO), isosorbide (1,4:3,6-dianhidro-D-sorbitol) (IS), and 1,4-cyclohexane dicaboxylic acid (CHDA), to enhance the thermal properties of polyesters. We have studied the effect of alicyclic monomers on the thermal properties, especially, on the glass transition temperature (T g ) of the amorphous copolyesters. On incorporating alicyclic diol monomers into poly(ethylene succinate-co-terephthalate) (PEST), T g of amorphous copolyesters was greatly increased. As the rigidity of alicyclic monomers increases, the effect becomes more significant. By comparing copolyesters synthesized with the same molar ratio of alicyclic monomers in the feed, we found that the molar ratio of IS in the copolyester product was much smaller than those of CHDM and CHDO. Nonetheless, IS induced the largest T g enhancement in copolyesters even though a much smaller molar ratio of IS was incorporated in the product compared to CHDM and CHDO. To study the effects of alicyclic diacid monomers, copolyesters containing CHDA were synthesized. As in the case of alicyclic diols, CHDA was found to be very effective in enhancing the thermal properties. Among these copolyesters produced, PEI30ST synthesized with 30 mol% of succinic acid (SA) and 70 mol% of dimethyl terephthalate (DMT) as diacid and 30 mol% of IS as diol exhibited the highest T g of 74.2 °C. And poly(ethylene-co-CHDM succinate-co-terephthalate) (PEC30ST70) and poly(ethylene-co-IS succinate-co-terephthalate) (PEI30ST70) exhibited high optical transmittance and were confirmed to be biodegradable, exhibiting about 50% and 40% biodegradability, respectively, after 50 days in a biodegradability test under compost conditions. Hence, these novel transparent biodegradable polyesters could be highly useful in flexible packaging applications requiring transparency.

Isoflurane has either neuroprotective or neurotoxic effects. High-dose oxygen is frequently used throughout the perioperative period. We hypothesized that hyperoxia will affect cell viability of rat pheochromocytoma (PC12) cells that were exposed to isoflurane and reactive oxygen species (ROS) may be involved. PC12 cells were exposed to 1.2% or 2.4% isoflurane for 6 or 24h respectively, and cell viability was evaluated. To investigate the effects of hyperoxia, PC12 cells were treated with 21%, 50%, or 95% oxygen and 2.4% isoflurane for 6h, and cell viability, TUNEL staining, ROS production, and expression of B-cell lymphoma 2 (BCL-2), BCL2-associated X protein (BAX), caspase-3 and beta-site APP cleaving enzyme (BACE) were measured. ROS involvement was evaluated using the ROS scavenger 2-mercaptopropiopylglycine (MPG). The viability of cells exposed to 2.4% isoflurane was lower than that of cells exposed to 1.2% isoflurane. Prolonged exposure (6h vs. 24h) to 2.4% isoflurane resulted in a profound reduction in cell viability. Treatment with 95% (but not 50%) oxygen enhanced the decrease in cell viability induced by 2.4% isoflurane alone. Levels of ROS, Bax, caspase-3 and BACE were increased, whereas expression of Bcl-2 was decreased, in cells treated with 95% oxygen plus 2.4% isoflurane compared with the control and 2.4% isoflurane plus air groups. MPG attenuated the effects of oxygen and isoflurane. In conclusion, isoflurane affects cell viability in a dose- and time-dependent manner. This effect is augmented by hyperoxia and may involve ROS, the mitochondrial apoptotic signaling pathway, and β-amyloid protein.

In this study, a computational chemistry methodology called as molecular modeling was been applied to explain several experiment results mechanistically. The reaction chosen for this study was to remove carbon dioxide, known as a primary greenhouse gas, by an epoxide via the carbon dioxide fixation to produce carbonates. This reaction inherently needs the use of catalysts because it has a significantly high activation barrier (55~59 kcal/mol). Among various types of catalysts, we studied in zeolitic imidazolate framework 90 (ZIF-90)/ionic liquid immobilized ZIF-90 (IL-ZIF-90), polystyrene-supported quaternized ammonium salt, KI/KI-glycine, and dimethylethanolamine (DMEA). First, probable reaction pathways were proposed based on calculated energetics by computational chemistry. The energetics was then used for the thermodynamic interpretation on the activity of catalysts. In the case of ZIF-90/IL-ZIF-90 and KI/KI-glycine, IL-ZIF-90 and KI-glycine showed better yields compared to their counterparts. The calculation proposed interesting results that it is not from the lowering of activation energy but from the unstable intermediates of ZIF-90 and KI-glycine. For DMEA, the calculated activation energy was ~42 kcal/mol, much lower than that of the non-catalytic reaction. A possible reaction pathway was located to confirm the interaction between -NH group from ammonium and oxygen from epoxide for polystyrene-supported quaternized ammonium salt.

Benzodiazepines and zolpidem are generally prescribed as sedative, hypnotics, anxiolytics or anticonvulsants. These drugs, however, are frequently misused in drug-facilitated crime. Therefore, a rapid and simple liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed for identification and quantification of benzodiazepines, zolpidem and their metabolites in urine using deuterium labeled internal standards (IS). Urine samples (120 μL) mixed with 80 μL of the IS solution were centrifuged. An aliquot (5 μL) of the sample solution was directly injected into the LC-MS/MS system for analysis. The mobile phases consisted of water and acetonitrile containing 2mM ammonium trifluoroacetate and 0.2% acetic acid. The analytical column was a Zorbax SB-C18 (100 mm × 2.1 mm i.d., 3.5 μm, Agilent). The separation and detection of 18 analytes were achieved within 10 min. Calibration curves were linear over the concentration ranges of 0.5-20 ng/mL (zolpidem), 1.0-40 ng/mL (flurazepam and temazepam), 2.5-100 ng/mL (7-aminoclonazepam, 1-hydroxymidazolam, midazolam, flunitrazepam and alprazolam), 5.0-200 ng/mL (zolpidem phenyl-4-carboxylic acid, α-hydroxyalprazolam, oxazepam, nordiazepam, triazolam, diazepam and α-hydroxytriazolam), 10-400 ng/mL (lorazepam and desalkylflurazepam) and 10-100 ng/mL (N-desmethylflunitrazepam) with the coefficients of determination (r(2)) above 0.9971. The dilution integrity of the analytes was examined for supplementation of short linear range. Dilution precision and accuracy were tested using two, four and ten-folds dilutions and they ranged from 3.7 to 14.4% and -12.8 to 12.5%, respectively. The process efficiency for this method was 63.0-104.6%. Intra- and inter-day precisions were less than 11.8% and 9.1%, while intra- and inter-day accuracies were less than -10.0 to 8.2%, respectively. The lower limits of quantification were lower than 10 ng/mL for each analyte. The applicability of the developed method was successfully verified with human urine samples from drug users (n=21). Direct urine sample injection and optimized mobile phases were introduced for simple sample preparation and high-sensitivity with the desired separation.

Summary: Exon junction complexes (EJCs) loaded onto spliced mRNAs during splicing serve as molecular markers for various post-transcriptional gene-regulatory processes, including nonsense-mediated mRNA decay (NMD). Although the composition and structure of EJCs are well characterized, the mechanism regulating EJC deposition remains unknown. Here we find that threonine 163 (T163) within the RNA-binding motif of eIF4A3 (a core EJC component) is phosphorylated by cyclin-dependent protein kinases 1 and 2 in a cell cycle-dependent manner. T163 phosphorylation hinders binding of eIF4A3 to spliced mRNAs and other EJC components. Instead, it promotes association of eIF4A3 with CWC22, which guides eIF4A3 to an active spliceosome. These molecular events ensure the fidelity of specific deposition of the EJC ∼20–24 nt upstream of an exon-exon junction. Accordingly, NMD is affected by T163 phosphorylation. Collectively, our data provide evidence that T163 phosphorylation affects EJC formation and, consequently, NMD efficiency in a cell cycle-dependent manner. : Ryu et al. show that eIF4A3 (a core EJC component) is phosphorylated at the threonine 163 position by CDK1 and CDK2 in a cell cycle-dependent manner. This event triggers EJC remodeling and affects NMD efficiency in a cell cycle-dependent manner. Keywords: exon junction complex, eIF4A3, nonsense-mediated mRNA decay, CDK, phosphorylation, cell cycle

Background/Aim: This study was designed to identify candidate proteins which can be used for visualization of pelvic autonomic nerve during nerve-sparing surgery. Materials and Methods: Both soft tissue and vesical branch of the inferior hypogastric nerve from five women were collected during surgery. These 10 tissue specimens were analysed using liquid chromatography–mass spectrometry (LC-MS) and Human Protein Atlas (HPA) for protein expression. The existence of nerve fibres was confirmed using haematoxylin and eosin (H&E) and anti-S-100 staining. Results: A total of 413 proteins were detected. There were three proteins (isoform 1 of fibronectin, protein S100-A8 and A9) which implied a relation with pelvic autonomic nerve. In nerve tissue from one case, the existence of nerve fibre was not confirmed. Conclusion: Further large studies are expected to present more nerve-specific candidate proteins which can be used for the easy and safe identification of autonomic nerves.

Periodontitis is a chronic inflammatory disease that contributes to the destruction of the gingiva. Porphyromonas gingivalis (P. gingivalis) can cause periodontitis via its pathogenic lipopolysaccharides (LPS). Melittin, a major component of bee venom, is known to have anti-inflammatory and antibacterial effects. However, the role of melittin in the inflammatory response has not been elucidated in periodontitis-like human keratinocytes. Therefore, we investigated the anti-inflammatory effects of melittin on a P. gingivalis LPS (PgLPS)-treated HaCaT human keratinocyte cell line. The cytotoxicity of melittin was measured using a human keratinocyte cell line, HaCaT, and a Cell Counting Kit-8. The effect of melittin on PgLPS-induced inflammation was determined with Western blot, real-time quantitative PCT, and immunofluorescence. PgLPS increased the expression of toll-like receptor (TLR) 4 and proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-8, and interferon-γ (IFN-γ). Moreover, PgLPS induced activation of the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), extracellular signal-regulated kinase (ERK), and protein kinase B/Akt. Melittin also inhibited the expression of proinflammatory cytokines by suppressing the activation of the NF-κB signaling pathway, ERK, and Akt. Melittin attenuates the PgLPS-induced inflammatory response and could therefore be applied in the treatment of periodontitis for anti-inflammatory effects.

Hp1404, identified from the venom of the scorpion Heterometrus petersii, displays antimicrobial activity with cytotoxicity. Several synthetic peptides were designed based on the parent peptide Hp1404 to reduce cytotoxicity and improve activity (deletion of glycine and phenylalanine, substitution with leucine and lysine). The analogue peptides generated comprised 12 amino acids and displayed amphipathic α-helical structures, with higher hydrophobic moments and net positive charge than those of the Hp1404. The analogues showed less hemolytic and toxic effects toward mammalian cells than the Hp1404, especially Hp1404-T1e, which exhibited particularly potent antibacterial and antibiofilm activities against multidrug-resistant Pseudomonas aeruginosa (MRPA) strains. The analogue peptide Hp1404-T1e was more stable against salt and trypsin than the Hp1404. Hp1404’s mechanism of action involves binding to lipopolysaccharide (LPS), thereby killing bacteria through membrane disruption. Hp1404-T1e kills bacteria more rapidly than Hp1404 and not only seems to bind more strongly to LPS but may also be able to enter bacterial cells and interact with their DNA. Additionally, Hp1404-T1e can effectively kill bacteria in vivo. The results of this study indicate that Hp1404-T1e not only displays antimicrobial activity, but is also functional in physiological conditions, confirming its potential use as an effective therapeutic agent against MRPA.

The complexation of boric acid (B(OH)3), the primary form of aqueous boron at moderate pH, with polyols is proposed and mechanistically studied as an efficient way to improve membrane processes such as reverse osmosis (RO) for removing boron in seawater by increasing the size of aqueous boron compounds. Computational chemistry based on the density functional theory (DFT) was used to manifest the reaction pathways of the complexation of B(OH)3 with various polyols such as glycerol, xylitol, and mannitol. The reaction energies were calculated as −80.6, −98.1, and −87.2 kcal/mol for glycerol, xylitol, and mannitol, respectively, indicating that xylitol is the most thermodynamically favorable for the complexation with B(OH)3. Moreover, the 1: 2 molar ratio of B(OH)3 to polyol was found to be more favorable than the ratio of 1: 1 for the complexation. Meanwhile, latest lab-scale actual RO experiments successfully supported our computational prediction that 2 moles of xylitol are the most effective as the complexing agent for 1 mole of B(OH)3 in aqueous solution.

The efficacy of a new chitosan (CS) based functional gel (FG), comprising multiwall carbon nanotube (MWNT)–poly(acrylic acid) (PAA)–poly(4-amino diphenyl amine) (PADPA), was investigated towards removal of hexavalent chromium (Cr(VI)). The new FG (CS–MWNT–PAA–PADPA/FG) was synthesized by free radical polymerization and cross-linking reactions. Characterization of the gel was done by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis, X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy. Pore parameters of the gel were evaluated by mercury porosimetry. Batch adsorption experiments were conducted to evaluate the FG for the removal of Cr(VI) from synthetic as well as field samples. The maximum Cr(VI) adsorption capacity was achieved for CS–MWNT–PAA–PADPA/FG owing to the synergistic effects from the individual components. The individual component such as (MWNT, CS, PAA and PADPA) in CS–MWNT–PAA–PADPA/FG plays a vital role to the improvement in Cr(VI) adsorption/removal. CS and PADPA were the nucleus for Cr(VI) adsorption through binding with –NH2 sites. PAA provides internal H+ ions within the gel and increases the adsorption efficiency of CS–MWNT–PAA–PADPA/FG over a wider pHs. PADPA provides additional –NH2 sites over CS for Cr(VI) adsorption. CS and PAA endow three dimensional hydrogel framework and controls the swelling/shrinking action of the hydrogel. MWNTs impart stability to the gel during adsorption–desorption cycle and improves re-useability. The Cr(VI) removal mechanism was investigated by FTIR spectroscopy, X-ray absorption spectroscopy and XPS. The results suggest complexation interactions between the multiple organic functional groups in FG and Cr(VI) and transformation of Cr(VI) to Cr(III). Furthermore, the new CS–MWNT–PAA–PADPA/FG is stable and recyclable, retaining about 85% of the removal efficiency up to three adsorption cycles.

Gene therapy using RNA interference can be directed against tumors through various strategies, but has been hindered owing to the inefficiency of non-viral delivery. To evaluate the antitumor effects of adenine nucleotide translocase-2 (ANT2) short hairpin RNA (shRNA) by intraperitoneal injection using the polyethylenimine (PEI) and an ultrasound gene delivery method, human breast carcinoma MDA-MB-231 cells were injected subcutaneously into NOG (NOD/Shi-scid/IL-2Rγ(null)) mice. The results showed greater tumor regression (*P

The abuse of antibiotics for disease treatment has led to the emergence of multidrug resistant bacteria. Antimicrobial peptides, found naturally in various organisms, have received increasing interest as alternatives to conventional antibiotics because of their broad spectrum antimicrobial activity and low cytotoxicity. In a previous report, Macropin, isolated from bee venom, exhibited antimicrobial activity against both gram-positive and negative bacteria. In the present study, Macropin was synthesized and its antibacterial and anti-biofilm activities were tested against bacterial strains, including gram-positive and negative bacteria, and drug resistant bacteria. Moreover, Macropin did not exhibit hemolytic activity and cytotoxicity to keratinocytes, whereas Melittin, as a positive control, showed very high toxicity. Circular dichroism assays showed that Macropin has an α-helical structure in membrane mimic environments. Macropin binds to peptidoglycan and lipopolysaccharide and kills the bacteria by disrupting their membranes. Moreover, the fractional inhibitory concentration index indicated that Macropin has additive and partially synergistic effects with conventional antibiotics against drug resistant bacteria. Thus, our study suggested that Macropin has potential for use of an antimicrobial agent for infectious bacteria, including drug resistant bacteria.

Mutations affecting RNA splicing factors are the most common genetic alterations in myelodysplastic syndrome (MDS) patients and occur in a mutually exclusive manner. The basis for the mutual exclusivity of these mutations and how they contribute to MDS is not well understood. Here we report that although different spliceosome gene mutations impart distinct effects on splicing, they are negatively selected for when co-expressed due to aberrant splicing and downregulation of regulators of hematopoietic stem cell survival and quiescence. In addition to this synthetic lethal interaction, mutations in the splicing factors SF3B1 and SRSF2 share convergent effects on aberrant splicing of mRNAs that promote nuclear factor κB signaling. These data identify shared consequences of splicing-factor mutations and the basis for their mutual exclusivity.

The G12 family of G protein alpha subunits has been shown to participate in the regulation of various physiological processes. However, the role of Gα12 in bone physiology has not been well described. Here, by micro‐CT analysis, we discovered that Gα12‐knockout mice have an osteopetrotic phenotype. Histological examination showed lower osteoclast number in femoral tissue of Gα12‐knockout mice compared to wild‐type mice. Additionally, in vitro osteoclastic differentiation of precursor cells with receptor activator of nuclear factor‐κB ligand (RANKL) showed that Gα12 deficiency decreased the number of osteoclast generated and the bone resorption activity. The induction of nuclear factor of activated T‐cell c1 (NFATc1), the key transcription factor of osteoclastogenesis, and the activation of RhoA by RANKL was also significantly suppressed by Gα12 deficiency. We further found that the RANKL induction of NFATc1 was not dependent on RhoA signalling, while osteoclast precursor migration and bone resorption required RhoA in the Gα12‐mediated regulation of osteoclasts. Therefore, Gα12 plays a role in differentiation through NFATc1 and in cell migration and resorption activity through RhoA during osteoclastogenesis.

Background Atopic dermatitis (AD) is identified by an increase in infiltrations of several inflammatory cells including type 2 helper (Th2) lymphocytes. Th2-related chemokines such as thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22), and pro-inflammatory cytokines including interleukin (IL)-1β and IL-6 are considered to play a crucial role in AD. Tumor necrosis factor (TNF)-α- and interferon (IFN)-γ induce the inflammatory condition through production of TARC, MDC, IL-1β and IL-6, and activations of related transcription factors, such as nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and signal transducer and activator of transcription (STAT) in keratinocytes. Apamin, a peptide component of bee venom, has been reported its beneficial activities in various diseases. However, anti-inflammatory effects of apamin on inflammatory condition in keratinocytes have not been explored. Therefore, the present study aimed to demonstrate the anti-inflammatory effect of apamin on TNF-α- and IFN-γ-induced inflammatory condition in keratinocytes. Methods HaCaT was used as human keratinocytes cell line. Cell Counting Kit-8 was performed to measure a cytotoxicity of apamin. The effects of apamin on TNF-α-/IFN-γ-induced inflammatory condition were determined by real-time PCR and Western blot analysis. Further, NF-κB signaling pathways, STAT1, and STAT3 were analyzed by Western blot and immunofluorescence. Results Apamin ameliorated the inflammatory condition through suppression of Th2-related chemokines and pro-inflammatory cytokines. Further, apamin down-regulated the activations of NF-κB signaling pathways and STATs in HaCaT cells. Conclusions These results suggest that apamin has therapeutic effect on AD through improvement of inflammatory condition.

The inhibitor NU 2058 [6-(cyclohexylmethoxy)-9H-purin-2-amine] leads to G1-phase cell cycle arrest in the marine diatom, Phaeodactylum tricornutum, by binding to two cyclin-dependent kinases, CDKA1 and CDKA2. NU 2058 has no effect on photosynthetic attributes, such as F v /F m , chlorophyll a/cell, levels of D2 PSII subunits, or RbcL; however, cell cycle arrest leads to unbalanced growth whereby photosynthetic products that can no longer be used for cell division are redirected toward carbohydrates and triacylglycerols (TAGs). Arrested cells up-regulate most genes involved in fatty acid synthesis, including acetyl-CoA carboxylase, and three out of five putative type II diglyceride acyltransferases (DGATs), the enzymes that catalyze TAG production. Correlation of transcriptomes in arrested cells with a flux balance model for P. tricornutum predicts that reactions in the mitochondrion that supply glycerate may support TAG synthesis. Our results reveal that sources of intermediate metabolites and macromolecular sinks are tightly coupled to the cell cycle in a marine diatom, and that arresting cells in the G1 phase leads to remodeling of intermediate metabolism and unbalanced growth. Refereed/Peer-reviewed

Constraint-based modeling has been shown, in many instances, to be useful for metabolic engineering by allowing the prediction of the metabolic phenotype resulting from genetic manipulations. But the basic premise of constraint-based modeling—that of applying constraints to preclude certain behaviors—only makes sense for certain genetic manipulations (such as knockouts and knockdowns). In particular, when genes (such as those associated with a heterologous pathway) are introduced under artificial control, it is unclear how to predict the correct behavior. In this paper, we introduce a modeling method that we call proportional flux forcing (PFF) to model artificially induced enzymatic genes. The model modifications introduced by PFF can be transformed into a set of simple mass balance constraints, which allows computational methods for strain optimization based on flux balance analysis (FBA) to be utilized. We applied PFF to the metabolic engineering of Escherichia coli (E. coli) for free fatty acid (FFA) production—a metabolic engineering problem that has attracted significant attention because FFAs are a precursor to liquid transportation fuels such as biodiesel and biogasoline. We show that PFF used in conjunction with FBA-based computational strain optimization methods can yield non-obvious genetic manipulation strategies that significantly increase FFA production in E. coli. The two mutant strains constructed and successfully tested in this work had peak fatty acid (FA) yields of 0.050 g FA/g carbon source (17.4% theoretical yield) and 0.035 g FA/g carbon source (12.3% theoretical yield) when they were grown using a mixed carbon source of glucose and casamino acids in a ratio of 2-to-1. These yields represent increases of 5.4- and 3.8-fold, respectively, over the baseline strain. Biotechnol. Bioeng. 2014;111: 2056–2066. © 2014 Wiley Periodicals, Inc.

Ovarian cancer (OVC) is one of the most difficult types of cancer to detect in the early stages of its development. There have been numerous attempts to identify a biomarker for OVC; however, an accurate diagnostic marker has yet to be identified. The present study profiled OVC candidate metabolites from the serum to identify potential diagnostic markers for OVC. Data regarding low-mass ions (LMIs) in the serum were obtained using matrix-assisted laser desorption/ionization (MALDI)-time-of-flight analysis. MALDI-mass spectrometry (MS) analysis of each serum sample was repeated six times in order to reduce the likelihood of experimental errors. The intensity of the LMI mass peaks were normalized using total peak area sums. The normalized intensity of LMI was used in principal component analysis-discriminant analysis to differentiate between 142 patients with OVC and 100 healthy control participants. Liquid chromatography-MS/MS was used to identify the selected LMIs. Extracted ion chromatogram analysis was used to measure the relative quantity of candidate metabolites from the LMI mass peak areas. The concentration of common metabolites in the serum was determined using ELISA. The top 20 LMI mass peaks with a weigh factor over 0.05 were selected to distinguish between the patients with OVC and the controls. Among the LMIs, two with 184.05 and 496.30 m/z were identified as L-homocysteic acid (HCA) and lysophosphatidylcholine (LPC) (16:0), respectively. The relative quantity of LPC (16:0) was found to be decreased in the OVC serum (P=0.05), while the quantity of HCA was observed to be significantly higher in the OVC serum (P

A rechargeable Li–air battery was studied using various carbon-supported metal-alloy catalysts as an air electrode. Carbon-supported Pt–Pd, Pd–Ir and Pt–Ru catalysts were prepared by an impregnation–reduction method. For comparison, carbon-supported Pt (Pt/C), Pd (Pd/C), Ir (Ir/C), and Ru (Ru/C) catalysts were also prepared by the same method; physicochemical characterizations were carried out using X-ray diffraction and transmission electron microscopy. The prepared catalysts were subsequently cast onto both nickel foam and GDL to be evaluated as air cathodes in Li–air batteries. Charge–discharge tests were carried out using a Swagelok-type cell, with both 1 M LiPF6 in PC:EC:DEC (3:2:5) and 1 M LiPF6 in TEGDME used as an electrolyte. Cycling performance and impedance tests were also conducted. The initial charge–discharge tests showed that alloying two different metals is an effective technique for decreasing the charging voltage. Among all electrodes tested, the Pt–Ru/C electrode was shown to possess a stable cycle performance and the highest round-trip efficiency.

This study provided sorption properties of chlorofluorocarbons (CFCs), and elucidated potential application of CFC sorption data in hydrogeochemistry. Prior sorption studies were reviewed for hydrophobic organic compounds similar to the CFCs, because there were only few CFC sorption studies. The CFCs are regarded as relatively conservative chemicals in groundwater environments based on their moderate hydrophobicity. However, thermally altered carbonaceous matter (TACM) can significantly increase sorption capacity and nonlinearity for hydrophobic organic compounds such as CFCs, compared to general soil organic matter. CFC sorption behavior are close to the sorption for reviewed organic chemicals. Therefore, the CFC sorption data can be used for determining hydrogeochemical properties and predicting transport of organic contaminants in TACM-containing aquifer environments.

Purpose The aim was to investigate the agreement for detection of progression of glaucomatous retinal nerve fibre layer defects (RNFLD) between optical coherence tomography-guided progression analysis (OCT GPA) and conventional red-free fundus photography. Methods Four hundred and fifteen glaucomatous eyes that underwent at least four serial red-free photographic and OCT examinations were included in the study. Based on the inspection of the red-free fundus photographs and GPA maps, RNFLD progression was defined as the development of a new defect, widening or deepening of a pre-existing RNFLD in red-free fundus photography (photographic progression) or ‘Likely Loss’ on a GPA map (GPA progression). The agreement of photographic and OCT GPA progression and the factors influencing it, including refractive error, severity of glaucoma (mean deviation of the visual field), type of RNFLD (localised versus diffuse), width of the baseline RNFLD, type of RNFLD progression (new defect, widening, deepening) and location of RNFLD progression (clock-hour sector) were assessed. Results Among the 415 eyes, 82 (19.8 per cent) showed photographic or GPA progression. Among the 82 eyes with progression, progression was detected only in red-free fundus photography in nine (11.0 per cent) eyes and only in GPA in 32 (39.0 per cent) eyes. In 41 eyes (50.0 per cent), progression was detected with both methods. Detection of RNFLD progression only in GPA was associated with a higher myopia, diffuse RNFLD, deepening of the RNFLD and RNFLD progression at the 6, 9 and 12 o'clock positions (p